Gene expression of cell diferenciation related factors of respiratory epithelium and its roll in the mucosal pathology of rhino sinusitis

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Genetic investigation : Several bands that appeared on the collagen gel following floating were extracted by the differential display method. The protein volume expression of 5 of the 25 sequenced bands was determined before and after floating by Northern blotting. Protein expression did not differ between the two measurement period. This lack of difference was attributed to the high frequency of false positive results using Northern blotting.Morphological confirmation of cell differentiation : Expression of cytokeratin the cultured cells was examined chronologically by an immunohistochemical technique using anti-cytokeratin 8, 13 and 14 monochronal antibodies, although the cultured cells showed no positive findings before floating, this analysis showed a gradual increase in positive staining after floating in the following pattern ; anti-cytokeratin 8 was positively stained from the middle to apical layer of the cultured cells and anti-cytokeratin 13 was positively stained only in the middle layer and anti-cytokeratin 14 was positively stained in the basal layer.Detection of ciliogenesis-specific substance : Several monochronal antibodies were produced, of which RPCS3a, the most eligible candidate specific to ciliogenesis, was characterized. RPCS3a was expressed in the cultured cell from the 3rd day after floating according to cellular ELISA.On the 10th day, RPCS3a was expressed most strongly, and many cultured cells contained secretory vesicles at their apical site as well as numerous centrioles indicating to be preciliated cells. ELISA revealed that RPCS3a is specific to the a sort of fucosylated carbohydrate H type 3. The expressed volume of H type 3 recognized by RPCS3a appears to reflect the stage of ciliogenesis. Based on the present data, we consider H type 3 to be a relativity specific markers of ciliogenesis and thus may provide a useful tool to clarify the mechanisms of ciliogenesis.